Target (protein/gene name):
dihydrofolate reductase-thymidylate synthase NCBI Gene # or RefSeq#:
Tc00.1047053509153.90 Gene ID:
3547029 Protein ID (NP or XP #) or Wolbachia#:
XP_815327.1 Organism:
Trypanosoma cruzi strain CL Brener Etiologic Risk Group (see link below):
Risk Group 2 (RG2) - Parasitic Agents Background/Disease Information:
Chagas disease is a tropical parasitic disease caused by the flagellate protozoan Trypanosoma cruzi. T. cruzi is commonly transmitted to humans and other mammals by an insect vector, the blood-sucking "kissing bugs" of the subfamily Triatominae (family Reduviidae) most commonly species belonging to the Triatoma, Rhodnius, and Panstrongylus genera.
The disease may also be spread through blood transfusion and organ transplantation, ingestion of food contaminated with parasites, and from a mother to her fetus.
The symptoms of Chagas disease vary over the course of an infection. In the early, acute stage, symptoms are mild and usually produce no more than local swelling at the site of infection. The initial acute phase is responsive to antiparasitic treatments, with 60–90% cure rates. After 4–8 weeks, individuals with active infections enter the chronic phase of Chagas disease that is asymptomatic for 60–80% of chronically infected individuals through their lifetime.
The antiparasitic treatments also appear to delay or prevent the development of disease symptoms during the chronic phase of the disease, but 20–40% of chronically infected individuals will still eventually develop life-threatening heart and digestive system disorders. The currently available antiparasitic treatments for Chagas disease are benznidazole and nifurtimox, which can cause temporary side effects in many patients including skin disorders, brain toxicity, and digestive system irritation.
Chagas disease is contracted primarily in the Americas, particularly in poor, rural areas of Mexico, Central America, and South America; very rarely, the disease has originated in the Southern United States. The insects that spread the disease are known by various local names, including vinchuca in Argentina, Bolivia, Chile and Paraguay, barbeiro (the barber) in Brazil, pito in Colombia, chinche in Central America, chipo in Venezuela, chupança, chinchorro, and "the kissing bug".
It is estimated that as many as 8 to 11 million people in Mexico, Central America, and South America have Chagas disease, most of whom do not know they are infected. Large-scale population movements from rural to urban areas of Latin America and to other regions of the world have increased the geographic distribution of Chagas disease, and cases have been noted in many countries, particularly in Europe. Control strategies have mostly focused on eliminating the triatomine insect vector and preventing transmission from other sources.
--Wikipedia Essentiality of this protein:
significant loss of fitness in bloodstream forms (6 days) in a study on African trypanosome [alsford] Complex of enzymes:
No EC#:
1.5.1.3 & 2.1.1.45 Link to BRENDA EC# page: http://www.brenda-enzymes.org/php/result_flat.php4?ecno=1.5.1.3 http://www.brenda-enzymes.org/php/result_flat.php4?ecno=2.1.1.45 Enzyme Assay information (spectrophotometric, coupled assay ?, reagents): -- link to Sigma (or other company) page for assay or assay reagents (substrates)
DHFR reaction diagram dihydrofolate NADPH
TS reaction diagram 5,10-methylenetetrahydrofolate dUMP -- link to paper that contains assay information
Assay for Dihydrofolate Reductase (1.5.1.3). http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/learning-center/assay-library/ec-number.html
An enzyme with this EC number or name (thymidylate synthase) or sequence has been assayed in Trypanosoma cruzi. http://www.brenda-enzymes.info/literature/lit.php4?e=2.1.1.45&r=485374 Structure Available (PDB or Homology model) -- PDB # or closest PDB entry if using homology model:
3INV (direct match) -- Query Coverage (if not direct match): -- Max Ident (if not direct match): Druggable Target (see Databases for this):
0.8 Current Inhibitors:
this paper reports potential inhibitors with IC50 values in the nanomolar range, the best around 20-30 nM.
the inhibitors synthesized for the paper were also active for human DHFR, and modifications were not effective in biasing the inhibition towards the parasite.
also refer to this paper. Expression Information (has it been expressed in bacterial cells):
Escherichia coli Bl21(de3) Plyss Plasmid pET11c Purification Method:
N/A Image of protein (PyMol or etc): http://www.pdb.org/pdb/images/3inv_bio_r_500.jpg Amino Acid Sequence: >gi|71653378|ref|XP_815327.1| dihydrofolate reductase-thymidylate synthase [Trypanosoma cruzi strain CL Brener] MSLFKIRMPETVAEGTRLALRAFSLVVAVDEHGGIGDGRSIPWNVPEDMKFFRDLTTKLRGKNVKPSPAK RNAVVMGRKTWDSIPPKFRPLPGRLNVVLSSTLTTQHLLDGLPDEEKRNLHADSIVAVNGGLEQALRLLA SPNYTPSIETVYCIGGGSVYAEALRPPCVHLLQAIYRTTIRASESSCSVFFRVPESGTEAAAGIEWQRET ISEELTSANGNETKYYFEKLIPRNREEEQYLSLVDRIIREGNVKHDRTGVGTLSIFGAQMRFSLRNNRLP LLTTKRVFWRGVCEELLWFLRGETYAKKLSDKGVHIWDDNGSRAFLDSRGLTEYEEMDLGPVYGFQWRHF GAAYTHHDANYDGQGVDQIKAIVETLKTNPDDRRMLFTAWNPSALPRMALPPCHLLAQFYVSNGELSCML YQRSCDMGLGVPFNIASYALLTILIAKATGLRPGELVHTLGDAHVYSNHVEPCNEQLKRVPRAFPYLVFR REREFLEDYEEGDMEVIDYAPYPPISMKMAV length of your protein in Amino Acids
521 Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website
58853.2 Da Molar Extinction coefficient of your protein at 280 nm wavelength:
73800 (reduced cysteines) CDS Gene Sequence: >gi|164451099:234741-236306 Trypanosoma cruzi strain CL Brener 1047053517157 genomic scaffold, whole genome shotgun sequence ATGTCGCTGTTTAAGATCCGCATGCCGGAGACGGTGGCGGAGGGGACACGTCTCGCACTGCGTGCCTTTT CCCTCGTTGTAGCGGTCGACGAGCACGGCGGCATTGGGGACGGGCGGTCGATCCCATGGAATGTGCCCGA GGACATGAAGTTTTTCCGGGATTTGACAACAAAACTGCGGGGGAAGAACGTCAAACCATCGCCTGCGAAG CGCAACGCCGTTGTGATGGGCCGCAAGACGTGGGACAGCATTCCGCCCAAATTCCGCCCGCTTCCCGGTC GCCTGAACGTCGTTTTGTCGTCCACCCTCACGACGCAACACCTCTTGGACGGCCTGCCAGACGAAGAAAA ACGGAATTTGCATGCCGACAGCATTGTGGCGGTAAATGGTGGACTCGAGCAGGCATTGCGGTTGCTCGCG TCCCCCAATTACACGCCGAGCATTGAGACGGTCTACTGCATTGGTGGCGGCTCTGTGTATGCGGAGGCCC TTCGCCCGCCGTGCGTTCATTTGCTGCAAGCCATTTACCGCACCACCATCCGCGCCAGCGAGTCTTCTTG CAGCGTGTTTTTTCGCGTTCCCGAGTCGGGTACGGAGGCGGCGGCGGGGATTGAGTGGCAGCGGGAGACA ATCTCCGAGGAGCTCACGTCGGCAAACGGCAACGAGACAAAGTACTACTTTGAGAAGCTCATCCCACGCA ACAGAGAGGAGGAGCAGTATCTAAGCCTTGTGGATCGCATCATACGCGAGGGGAATGTGAAGCATGACCG CACTGGGGTGGGCACGCTCTCCATCTTTGGCGCACAGATGCGTTTCTCGCTCCGCAACAATCGCCTACCA CTTTTGACAACGAAGCGGGTTTTTTGGCGCGGTGTGTGCGAGGAACTGCTGTGGTTTCTACGGGGCGAAA CATATGCCAAAAAGTTAAGTGACAAGGGAGTCCACATATGGGACGACAACGGCTCGCGCGCGTTTCTTGA CAGTCGTGGCCTCACAGAGTACGAGGAGATGGACCTCGGTCCTGTCTACGGCTTTCAGTGGCGTCACTTT GGTGCCGCCTACACACACCATGATGCCAACTATGACGGACAGGGTGTGGATCAAATCAAGGCAATCGTTG AAACGCTAAAGACAAATCCTGACGATCGCCGGATGCTATTCACGGCATGGAATCCGAGCGCTTTGCCCAG GATGGCCCTACCACCGTGCCACCTGCTCGCACAGTTTTACGTGAGCAATGGGGAACTGTCTTGCATGCTC TACCAGCGCTCATGTGACATGGGCCTTGGCGTCCCGTTCAACATTGCCTCCTACGCCCTCTTGACCATCC TCATCGCAAAGGCGACGGGGCTGCGGCCGGGAGAGTTGGTGCACACCCTTGGTGACGCTCACGTGTACAG CAATCACGTCGAGCCGTGCAATGAGCAACTGAAACGTGTGCCACGTGCCTTTCCGTATCTTGTGTTTCGC CGCGAGCGCGAGTTCCTGGAGGACTACGAGGAGGGTGACATGGAGGTCATTGACTATGCGCCATACCCTC CCATCTCCATGAAGATGGCGGTTTAG GC% Content for gene:
57.54% CDS Gene Sequence (codon optimized) - after having done Primer Design: ATGTCCCTGTTCAAAATCCGTATGCCTGAGACTGTGGCAGAAGGCACCCGCCTGGCGCTC CGTGCGTTTTCTCTCGTTGTGGCCGTTGATGAGCACGGTGGTATCGGTGATGGCCGCTCC ATCCCTTGGAACGTACCGGAGGACATGAAATTCTTCCGTGACCTGACGACCAAACTCCGT GGTAAAAACGTTAAACCATCTCCGGCGAAGCGTAACGCAGTTGTTATGGGTCGTAAAACC TGGGATTCTATCCCGCCGAAATTCCGTCCTCTGCCAGGCCGCCTGAATGTTGTTCTGTCC TCTACTCTGACCACCCAGCATCTCCTCGACGGTCTCCCGGATGAAGAGAAACGTAACCTG CACGCGGACTCTATCGTTGCGGTTAACGGCGGTCTGGAACAGGCCCTGCGCCTGCTCGCC TCTCCAAACTATACCCCGTCTATCGAAACTGTGTACTGCATCGGCGGTGGTTCTGTTTAT GCTGAAGCGCTGCGTCCGCCATGCGTTCATCTGCTGCAGGCGATCTACCGTACCACCATC CGTGCCTCTGAGTCTAGCTGTAGCGTGTTCTTTCGTGTCCCGGAATCTGGTACCGAAGCT GCCGCAGGTATCGAATGGCAGCGTGAGACTATTTCCGAGGAACTGACCTCTGCGAACGGT AACGAAACCAAATACTACTTCGAAAAACTGATCCCGCGCAACCGTGAGGAAGAACAATAC CTGTCTCTGGTTGACCGTATCATTCGTGAAGGTAACGTCAAACATGACCGCACTGGTGTT GGTACCCTCTCTATTTTCGGTGCGCAGATGCGCTTCTCTCTCCGTAATAACCGCCTGCCA CTGCTCACCACCAAACGTGTTTTCTGGCGTGGTGTTTGCGAAGAACTGCTGTGGTTCCTG CGTGGCGAAACCTACGCGAAAAAACTCTCTGACAAAGGTGTCCACATCTGGGACGACAAC GGTTCTCGCGCGTTCCTGGACAGCCGTGGCCTGACCGAATACGAAGAAATGGACCTGGGT CCTGTTTACGGCTTTCAGTGGCGTCACTTTGGTGCTGCGTACACCCACCACGACGCGAAC TACGACGGCCAAGGTGTTGACCAGATCAAAGCGATCGTTGAAACCCTGAAAACGAATCCA GATGATCGTCGTATGCTGTTCACTGCTTGGAATCCGTCTGCCCTGCCGCGTATGGCTCTC CCTCCTTGCCACCTGCTGGCGCAATTCTACGTGTCTAACGGTGAGCTGTCTTGCATGCTG TACCAGCGTAGCTGCGACATGGGTCTGGGTGTTCCATTCAACATCGCTTCTTACGCCCTC CTGACCATCCTGATCGCGAAAGCGACCGGCCTCCGCCCAGGTGAACTGGTTCATACCCTG GGTGACGCGCACGTTTACTCTAACCACGTTGAACCGTGCAACGAACAACTCAAACGCGTA CCTCGTGCCTTTCCGTACCTGGTTTTCCGTCGTGAACGTGAATTTCTGGAAGACTATGAA GAAGGTGACATGGAAGTTATCGACTATGCGCCGTACCCACCTATCTCTATGAAAATGGCG GTTTAA GC% Content for gene (codon optimized):
54.15% Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~60 nt long primers): 1 ATGTCCCTGTTCAAAATCCGTATGCCTGAGACTGTGGCAGAAGGCACCCGCCT 53 2 CGTGCTCATCAACGGCCACAACGAGAGAAAACGCACGGAGCGCCAGGCGGGTGCCTTCTG 60 3 GGCCGTTGATGAGCACGGTGGTATCGGTGATGGCCGCTCCATCCCTTGGAACGTACCGGA 60 4 CCACGGAGTTTGGTCGTCAGGTCACGGAAGAATTTCATGTCCTCCGGTACGTTCCAAGGG 60 5 GACGACCAAACTCCGTGGTAAAAACGTTAAACCATCTCCGGCGAAGCGTAACGCAGTTGT 60 6 GGAATTTCGGCGGGATAGAATCCCAGGTTTTACGACCCATAACAACTGCGTTACGCTTCG 60 7 ATTCTATCCCGCCGAAATTCCGTCCTCTGCCAGGCCGCCTGAATGTTGTTCTGTCCTCTA 60 8 TCCGGGAGACCGTCGAGGAGATGCTGGGTGGTCAGAGTAGAGGACAGAACAACATTCAGG 60 9 TCGACGGTCTCCCGGATGAAGAGAAACGTAACCTGCACGCGGACTCTATCGTTGCGGTTA 60 10 GGAGAGGCGAGCAGGCGCAGGGCCTGTTCCAGACCGCCGTTAACCGCAACGATAGAGTCC 60 11 GCCTGCTCGCCTCTCCAAACTATACCCCGTCTATCGAAACTGTGTACTGCATCGGCGGTG 60 12 AGATGAACGCATGGCGGACGCAGCGCTTCAGCATAAACAGAACCACCGCCGATGCAGTAC 60 13 CCGCCATGCGTTCATCTGCTGCAGGCGATCTACCGTACCACCATCCGTGCCTCTGAGTCT 60 14 TCGGTACCAGATTCCGGGACACGAAAGAACACGCTACAGCTAGACTCAGAGGCACGGATG 60 15 CCCGGAATCTGGTACCGAAGCTGCCGCAGGTATCGAATGGCAGCGTGAGACTATTTCCGA 60 16 GTAGTATTTGGTTTCGTTACCGTTCGCAGAGGTCAGTTCCTCGGAAATAGTCTCACGCTG 60 17 ACGGTAACGAAACCAAATACTACTTCGAAAAACTGATCCCGCGCAACCGTGAGGAAGAAC 60 18 GTTACCTTCACGAATGATACGGTCAACCAGAGACAGGTATTGTTCTTCCTCACGGTTGCG 60 19 CCGTATCATTCGTGAAGGTAACGTCAAACATGACCGCACTGGTGTTGGTACCCTCTCTAT 60 20 CGGTTATTACGGAGAGAGAAGCGCATCTGCGCACCGAAAATAGAGAGGGTACCAACACCA 60 21 GCTTCTCTCTCCGTAATAACCGCCTGCCACTGCTCACCACCAAACGTGTTTTCTGGCGTG 60 22 GTTTCGCCACGCAGGAACCACAGCAGTTCTTCGCAAACACCACGCCAGAAAACACGTTTG 60 23 TTCCTGCGTGGCGAAACCTACGCGAAAAAACTCTCTGACAAAGGTGTCCACATCTGGGAC 60 24 GTCAGGCCACGGCTGTCCAGGAACGCGCGAGAACCGTTGTCGTCCCAGATGTGGACACCT 60 25 ACAGCCGTGGCCTGACCGAATACGAAGAAATGGACCTGGGTCCTGTTTACGGCTTTCAGT 60 26 TCGCGTCGTGGTGGGTGTACGCAGCACCAAAGTGACGCCACTGAAAGCCGTAAACAGGAC 60 27 CCCACCACGACGCGAACTACGACGGCCAAGGTGTTGACCAGATCAAAGCGATCGTTGAAA 60 28 GAACAGCATACGACGATCATCTGGATTCGTTTTCAGGGTTTCAACGATCGCTTTGATCTG 60 29 GATGATCGTCGTATGCTGTTCACTGCTTGGAATCCGTCTGCCCTGCCGCGTATGGCTCTC 60 30 CACCGTTAGACACGTAGAATTGCGCCAGCAGGTGGCAAGGAGGGAGAGCCATACGCGGCA 60 31 CAATTCTACGTGTCTAACGGTGAGCTGTCTTGCATGCTGTACCAGCGTAGCTGCGACATG 60 32 CAGGAGGGCGTAAGAAGCGATGTTGAATGGAACACCCAGACCCATGTCGCAGCTACGCTG 60 33 GCTTCTTACGCCCTCCTGACCATCCTGATCGCGAAAGCGACCGGCCTCCGCCCAGGTGAA 60 34 ACGTGGTTAGAGTAAACGTGCGCGTCACCCAGGGTATGAACCAGTTCACCTGGGCGGAGG 60 35 GCACGTTTACTCTAACCACGTTGAACCGTGCAACGAACAACTCAAACGCGTACCTCGTGC 60 36 TTCCAGAAATTCACGTTCACGACGGAAAACCAGGTACGGAAAGGCACGAGGTACGCGTTT 60 37 CGTGAACGTGAATTTCTGGAAGACTATGAAGAAGGTGACATGGAAGTTATCGACTATGCG 60 38 TTAAACCGCCATTTTCATAGAGATAGGTGGGTACGGCGCATAGTCGATAACTTCCATG 58 DNA Works Output File: LOGFILE.txt Primer design results for 'tail' primers (this is just 2 sequences):
upstream: TACTTCCAATCCATGTCCCTGTTCAAAATCCGTA
downstream: TATCCACCTTTACTGTTAAACCGCCATTTTCATAGAGA codedata
http://www.ncbi.nlm.nih.gov/gene?term=Tc00.1047053509153.90
http://www.ncbi.nlm.nih.gov/protein/XP_815327.1
http://www.pdb.org/pdb/explore/explore.do?structureId=3INV
Target (protein/gene name):
dihydrofolate reductase-thymidylate synthase
NCBI Gene # or RefSeq#:
Tc00.1047053509153.90
Gene ID:
3547029
Protein ID (NP or XP #) or Wolbachia#:
XP_815327.1
Organism:
Trypanosoma cruzi strain CL Brener
Etiologic Risk Group (see link below):
Risk Group 2 (RG2) - Parasitic Agents
Background/Disease Information:
Chagas disease is a tropical parasitic disease caused by the flagellate protozoan Trypanosoma cruzi. T. cruzi is commonly transmitted to humans and other mammals by an insect vector, the blood-sucking "kissing bugs" of the subfamily Triatominae (family Reduviidae) most commonly species belonging to the Triatoma, Rhodnius, and Panstrongylus genera.
The disease may also be spread through blood transfusion and organ transplantation, ingestion of food contaminated with parasites, and from a mother to her fetus.
The symptoms of Chagas disease vary over the course of an infection. In the early, acute stage, symptoms are mild and usually produce no more than local swelling at the site of infection. The initial acute phase is responsive to antiparasitic treatments, with 60–90% cure rates. After 4–8 weeks, individuals with active infections enter the chronic phase of Chagas disease that is asymptomatic for 60–80% of chronically infected individuals through their lifetime.
The antiparasitic treatments also appear to delay or prevent the development of disease symptoms during the chronic phase of the disease, but 20–40% of chronically infected individuals will still eventually develop life-threatening heart and digestive system disorders. The currently available antiparasitic treatments for Chagas disease are benznidazole and nifurtimox, which can cause temporary side effects in many patients including skin disorders, brain toxicity, and digestive system irritation.
Chagas disease is contracted primarily in the Americas, particularly in poor, rural areas of Mexico, Central America, and South America; very rarely, the disease has originated in the Southern United States. The insects that spread the disease are known by various local names, including vinchuca in Argentina, Bolivia, Chile and Paraguay, barbeiro (the barber) in Brazil, pito in Colombia, chinche in Central America, chipo in Venezuela, chupança, chinchorro, and "the kissing bug".
It is estimated that as many as 8 to 11 million people in Mexico, Central America, and South America have Chagas disease, most of whom do not know they are infected. Large-scale population movements from rural to urban areas of Latin America and to other regions of the world have increased the geographic distribution of Chagas disease, and cases have been noted in many countries, particularly in Europe. Control strategies have mostly focused on eliminating the triatomine insect vector and preventing transmission from other sources.
--Wikipedia
Essentiality of this protein:
significant loss of fitness in bloodstream forms (6 days) in a study on African trypanosome [alsford]
Complex of enzymes:
No
EC#:
1.5.1.3 & 2.1.1.45
Link to BRENDA EC# page:
http://www.brenda-enzymes.org/php/result_flat.php4?ecno=1.5.1.3
http://www.brenda-enzymes.org/php/result_flat.php4?ecno=2.1.1.45
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
-- link to Sigma (or other company) page for assay or assay reagents (substrates)
DHFR
reaction diagram
dihydrofolate
NADPH
TS
reaction diagram
5,10-methylenetetrahydrofolate
dUMP
-- link to paper that contains assay information
Assay for Dihydrofolate Reductase (1.5.1.3).
http://www.sigmaaldrich.com/life-science/metabolomics/enzyme-explorer/learning-center/assay-library/ec-number.html
An enzyme with this EC number or name (thymidylate synthase) or sequence has been assayed in Trypanosoma cruzi.
http://www.brenda-enzymes.info/literature/lit.php4?e=2.1.1.45&r=485374
Structure Available (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model:
3INV (direct match)
-- Query Coverage (if not direct match):
-- Max Ident (if not direct match):
Druggable Target (see Databases for this):
0.8
Current Inhibitors:
this paper reports potential inhibitors with IC50 values in the nanomolar range, the best around 20-30 nM.
the inhibitors synthesized for the paper were also active for human DHFR, and modifications were not effective in biasing the inhibition towards the parasite.
also refer to this paper.
Expression Information (has it been expressed in bacterial cells):
Escherichia coli Bl21(de3) Plyss Plasmid pET11c
Purification Method:
N/A
Image of protein (PyMol or etc):
http://www.pdb.org/pdb/images/3inv_bio_r_500.jpg
Amino Acid Sequence:
>gi|71653378|ref|XP_815327.1| dihydrofolate reductase-thymidylate synthase [Trypanosoma cruzi strain CL Brener]
MSLFKIRMPETVAEGTRLALRAFSLVVAVDEHGGIGDGRSIPWNVPEDMKFFRDLTTKLRGKNVKPSPAK
RNAVVMGRKTWDSIPPKFRPLPGRLNVVLSSTLTTQHLLDGLPDEEKRNLHADSIVAVNGGLEQALRLLA
SPNYTPSIETVYCIGGGSVYAEALRPPCVHLLQAIYRTTIRASESSCSVFFRVPESGTEAAAGIEWQRET
ISEELTSANGNETKYYFEKLIPRNREEEQYLSLVDRIIREGNVKHDRTGVGTLSIFGAQMRFSLRNNRLP
LLTTKRVFWRGVCEELLWFLRGETYAKKLSDKGVHIWDDNGSRAFLDSRGLTEYEEMDLGPVYGFQWRHF
GAAYTHHDANYDGQGVDQIKAIVETLKTNPDDRRMLFTAWNPSALPRMALPPCHLLAQFYVSNGELSCML
YQRSCDMGLGVPFNIASYALLTILIAKATGLRPGELVHTLGDAHVYSNHVEPCNEQLKRVPRAFPYLVFR
REREFLEDYEEGDMEVIDYAPYPPISMKMAV
length of your protein in Amino Acids
521
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website
58853.2 Da
Molar Extinction coefficient of your protein at 280 nm wavelength:
73800 (reduced cysteines)
CDS Gene Sequence:
>gi|164451099:234741-236306 Trypanosoma cruzi strain CL Brener 1047053517157 genomic scaffold, whole genome shotgun sequence
ATGTCGCTGTTTAAGATCCGCATGCCGGAGACGGTGGCGGAGGGGACACGTCTCGCACTGCGTGCCTTTT
CCCTCGTTGTAGCGGTCGACGAGCACGGCGGCATTGGGGACGGGCGGTCGATCCCATGGAATGTGCCCGA
GGACATGAAGTTTTTCCGGGATTTGACAACAAAACTGCGGGGGAAGAACGTCAAACCATCGCCTGCGAAG
CGCAACGCCGTTGTGATGGGCCGCAAGACGTGGGACAGCATTCCGCCCAAATTCCGCCCGCTTCCCGGTC
GCCTGAACGTCGTTTTGTCGTCCACCCTCACGACGCAACACCTCTTGGACGGCCTGCCAGACGAAGAAAA
ACGGAATTTGCATGCCGACAGCATTGTGGCGGTAAATGGTGGACTCGAGCAGGCATTGCGGTTGCTCGCG
TCCCCCAATTACACGCCGAGCATTGAGACGGTCTACTGCATTGGTGGCGGCTCTGTGTATGCGGAGGCCC
TTCGCCCGCCGTGCGTTCATTTGCTGCAAGCCATTTACCGCACCACCATCCGCGCCAGCGAGTCTTCTTG
CAGCGTGTTTTTTCGCGTTCCCGAGTCGGGTACGGAGGCGGCGGCGGGGATTGAGTGGCAGCGGGAGACA
ATCTCCGAGGAGCTCACGTCGGCAAACGGCAACGAGACAAAGTACTACTTTGAGAAGCTCATCCCACGCA
ACAGAGAGGAGGAGCAGTATCTAAGCCTTGTGGATCGCATCATACGCGAGGGGAATGTGAAGCATGACCG
CACTGGGGTGGGCACGCTCTCCATCTTTGGCGCACAGATGCGTTTCTCGCTCCGCAACAATCGCCTACCA
CTTTTGACAACGAAGCGGGTTTTTTGGCGCGGTGTGTGCGAGGAACTGCTGTGGTTTCTACGGGGCGAAA
CATATGCCAAAAAGTTAAGTGACAAGGGAGTCCACATATGGGACGACAACGGCTCGCGCGCGTTTCTTGA
CAGTCGTGGCCTCACAGAGTACGAGGAGATGGACCTCGGTCCTGTCTACGGCTTTCAGTGGCGTCACTTT
GGTGCCGCCTACACACACCATGATGCCAACTATGACGGACAGGGTGTGGATCAAATCAAGGCAATCGTTG
AAACGCTAAAGACAAATCCTGACGATCGCCGGATGCTATTCACGGCATGGAATCCGAGCGCTTTGCCCAG
GATGGCCCTACCACCGTGCCACCTGCTCGCACAGTTTTACGTGAGCAATGGGGAACTGTCTTGCATGCTC
TACCAGCGCTCATGTGACATGGGCCTTGGCGTCCCGTTCAACATTGCCTCCTACGCCCTCTTGACCATCC
TCATCGCAAAGGCGACGGGGCTGCGGCCGGGAGAGTTGGTGCACACCCTTGGTGACGCTCACGTGTACAG
CAATCACGTCGAGCCGTGCAATGAGCAACTGAAACGTGTGCCACGTGCCTTTCCGTATCTTGTGTTTCGC
CGCGAGCGCGAGTTCCTGGAGGACTACGAGGAGGGTGACATGGAGGTCATTGACTATGCGCCATACCCTC
CCATCTCCATGAAGATGGCGGTTTAG
GC% Content for gene:
57.54%
CDS Gene Sequence (codon optimized) - after having done Primer Design:
ATGTCCCTGTTCAAAATCCGTATGCCTGAGACTGTGGCAGAAGGCACCCGCCTGGCGCTC
CGTGCGTTTTCTCTCGTTGTGGCCGTTGATGAGCACGGTGGTATCGGTGATGGCCGCTCC
ATCCCTTGGAACGTACCGGAGGACATGAAATTCTTCCGTGACCTGACGACCAAACTCCGT
GGTAAAAACGTTAAACCATCTCCGGCGAAGCGTAACGCAGTTGTTATGGGTCGTAAAACC
TGGGATTCTATCCCGCCGAAATTCCGTCCTCTGCCAGGCCGCCTGAATGTTGTTCTGTCC
TCTACTCTGACCACCCAGCATCTCCTCGACGGTCTCCCGGATGAAGAGAAACGTAACCTG
CACGCGGACTCTATCGTTGCGGTTAACGGCGGTCTGGAACAGGCCCTGCGCCTGCTCGCC
TCTCCAAACTATACCCCGTCTATCGAAACTGTGTACTGCATCGGCGGTGGTTCTGTTTAT
GCTGAAGCGCTGCGTCCGCCATGCGTTCATCTGCTGCAGGCGATCTACCGTACCACCATC
CGTGCCTCTGAGTCTAGCTGTAGCGTGTTCTTTCGTGTCCCGGAATCTGGTACCGAAGCT
GCCGCAGGTATCGAATGGCAGCGTGAGACTATTTCCGAGGAACTGACCTCTGCGAACGGT
AACGAAACCAAATACTACTTCGAAAAACTGATCCCGCGCAACCGTGAGGAAGAACAATAC
CTGTCTCTGGTTGACCGTATCATTCGTGAAGGTAACGTCAAACATGACCGCACTGGTGTT
GGTACCCTCTCTATTTTCGGTGCGCAGATGCGCTTCTCTCTCCGTAATAACCGCCTGCCA
CTGCTCACCACCAAACGTGTTTTCTGGCGTGGTGTTTGCGAAGAACTGCTGTGGTTCCTG
CGTGGCGAAACCTACGCGAAAAAACTCTCTGACAAAGGTGTCCACATCTGGGACGACAAC
GGTTCTCGCGCGTTCCTGGACAGCCGTGGCCTGACCGAATACGAAGAAATGGACCTGGGT
CCTGTTTACGGCTTTCAGTGGCGTCACTTTGGTGCTGCGTACACCCACCACGACGCGAAC
TACGACGGCCAAGGTGTTGACCAGATCAAAGCGATCGTTGAAACCCTGAAAACGAATCCA
GATGATCGTCGTATGCTGTTCACTGCTTGGAATCCGTCTGCCCTGCCGCGTATGGCTCTC
CCTCCTTGCCACCTGCTGGCGCAATTCTACGTGTCTAACGGTGAGCTGTCTTGCATGCTG
TACCAGCGTAGCTGCGACATGGGTCTGGGTGTTCCATTCAACATCGCTTCTTACGCCCTC
CTGACCATCCTGATCGCGAAAGCGACCGGCCTCCGCCCAGGTGAACTGGTTCATACCCTG
GGTGACGCGCACGTTTACTCTAACCACGTTGAACCGTGCAACGAACAACTCAAACGCGTA
CCTCGTGCCTTTCCGTACCTGGTTTTCCGTCGTGAACGTGAATTTCTGGAAGACTATGAA
GAAGGTGACATGGAAGTTATCGACTATGCGCCGTACCCACCTATCTCTATGAAAATGGCG
GTTTAA
GC% Content for gene (codon optimized):
54.15%
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~60 nt long primers):
1 ATGTCCCTGTTCAAAATCCGTATGCCTGAGACTGTGGCAGAAGGCACCCGCCT 53
2 CGTGCTCATCAACGGCCACAACGAGAGAAAACGCACGGAGCGCCAGGCGGGTGCCTTCTG 60
3 GGCCGTTGATGAGCACGGTGGTATCGGTGATGGCCGCTCCATCCCTTGGAACGTACCGGA 60
4 CCACGGAGTTTGGTCGTCAGGTCACGGAAGAATTTCATGTCCTCCGGTACGTTCCAAGGG 60
5 GACGACCAAACTCCGTGGTAAAAACGTTAAACCATCTCCGGCGAAGCGTAACGCAGTTGT 60
6 GGAATTTCGGCGGGATAGAATCCCAGGTTTTACGACCCATAACAACTGCGTTACGCTTCG 60
7 ATTCTATCCCGCCGAAATTCCGTCCTCTGCCAGGCCGCCTGAATGTTGTTCTGTCCTCTA 60
8 TCCGGGAGACCGTCGAGGAGATGCTGGGTGGTCAGAGTAGAGGACAGAACAACATTCAGG 60
9 TCGACGGTCTCCCGGATGAAGAGAAACGTAACCTGCACGCGGACTCTATCGTTGCGGTTA 60
10 GGAGAGGCGAGCAGGCGCAGGGCCTGTTCCAGACCGCCGTTAACCGCAACGATAGAGTCC 60
11 GCCTGCTCGCCTCTCCAAACTATACCCCGTCTATCGAAACTGTGTACTGCATCGGCGGTG 60
12 AGATGAACGCATGGCGGACGCAGCGCTTCAGCATAAACAGAACCACCGCCGATGCAGTAC 60
13 CCGCCATGCGTTCATCTGCTGCAGGCGATCTACCGTACCACCATCCGTGCCTCTGAGTCT 60
14 TCGGTACCAGATTCCGGGACACGAAAGAACACGCTACAGCTAGACTCAGAGGCACGGATG 60
15 CCCGGAATCTGGTACCGAAGCTGCCGCAGGTATCGAATGGCAGCGTGAGACTATTTCCGA 60
16 GTAGTATTTGGTTTCGTTACCGTTCGCAGAGGTCAGTTCCTCGGAAATAGTCTCACGCTG 60
17 ACGGTAACGAAACCAAATACTACTTCGAAAAACTGATCCCGCGCAACCGTGAGGAAGAAC 60
18 GTTACCTTCACGAATGATACGGTCAACCAGAGACAGGTATTGTTCTTCCTCACGGTTGCG 60
19 CCGTATCATTCGTGAAGGTAACGTCAAACATGACCGCACTGGTGTTGGTACCCTCTCTAT 60
20 CGGTTATTACGGAGAGAGAAGCGCATCTGCGCACCGAAAATAGAGAGGGTACCAACACCA 60
21 GCTTCTCTCTCCGTAATAACCGCCTGCCACTGCTCACCACCAAACGTGTTTTCTGGCGTG 60
22 GTTTCGCCACGCAGGAACCACAGCAGTTCTTCGCAAACACCACGCCAGAAAACACGTTTG 60
23 TTCCTGCGTGGCGAAACCTACGCGAAAAAACTCTCTGACAAAGGTGTCCACATCTGGGAC 60
24 GTCAGGCCACGGCTGTCCAGGAACGCGCGAGAACCGTTGTCGTCCCAGATGTGGACACCT 60
25 ACAGCCGTGGCCTGACCGAATACGAAGAAATGGACCTGGGTCCTGTTTACGGCTTTCAGT 60
26 TCGCGTCGTGGTGGGTGTACGCAGCACCAAAGTGACGCCACTGAAAGCCGTAAACAGGAC 60
27 CCCACCACGACGCGAACTACGACGGCCAAGGTGTTGACCAGATCAAAGCGATCGTTGAAA 60
28 GAACAGCATACGACGATCATCTGGATTCGTTTTCAGGGTTTCAACGATCGCTTTGATCTG 60
29 GATGATCGTCGTATGCTGTTCACTGCTTGGAATCCGTCTGCCCTGCCGCGTATGGCTCTC 60
30 CACCGTTAGACACGTAGAATTGCGCCAGCAGGTGGCAAGGAGGGAGAGCCATACGCGGCA 60
31 CAATTCTACGTGTCTAACGGTGAGCTGTCTTGCATGCTGTACCAGCGTAGCTGCGACATG 60
32 CAGGAGGGCGTAAGAAGCGATGTTGAATGGAACACCCAGACCCATGTCGCAGCTACGCTG 60
33 GCTTCTTACGCCCTCCTGACCATCCTGATCGCGAAAGCGACCGGCCTCCGCCCAGGTGAA 60
34 ACGTGGTTAGAGTAAACGTGCGCGTCACCCAGGGTATGAACCAGTTCACCTGGGCGGAGG 60
35 GCACGTTTACTCTAACCACGTTGAACCGTGCAACGAACAACTCAAACGCGTACCTCGTGC 60
36 TTCCAGAAATTCACGTTCACGACGGAAAACCAGGTACGGAAAGGCACGAGGTACGCGTTT 60
37 CGTGAACGTGAATTTCTGGAAGACTATGAAGAAGGTGACATGGAAGTTATCGACTATGCG 60
38 TTAAACCGCCATTTTCATAGAGATAGGTGGGTACGGCGCATAGTCGATAACTTCCATG 58
DNA Works Output File:
LOGFILE.txt
Primer design results for 'tail' primers (this is just 2 sequences):
upstream: TACTTCCAATCCATGTCCCTGTTCAAAATCCGTA
downstream: TATCCACCTTTACTGTTAAACCGCCATTTTCATAGAGA
code data